Abstract Resistance to multiple antibiotics has emerged in many bacterial pathogens, with life-threatening infections expected to increase to 10 million cases annually worldwide by 2050. Development of new chemotherapeutics is a high priority with several national/international programs beginning to evaluate non-antibiotic FDA-approved drugs for infectious disease indications. We responded to this new healthcare initiative by screening 780 FDA- approved drugs against a Tier-1 select agent in our unique biosafety containment facility. This approach is cost effective as it significantly shortens the time to market by leveraging well-defined drug structures and pharmacological properties, as well as safety profiles in patients. We identified three new ?drug leads? (doxapram, a breathing stimulant; amoxapine, an anti-depressant; and trifluoperazine, an anti-psychotic) that showed excellent protection from pneumonic plague in mice infected with Yersinia pestis. These drugs were efficacious when used as a treatment option administered one to three times at a much lower human equivalent doses. Our data show that these drugs also extend protection against an urgent healthcare threat Clostridium difficile, the leading cause of antibiotic-associated diarrhea, and multiple drug resistant (MDR) Salmonella Typhimurium, representing a serious public health threat. Whole transcriptome analysis of macrophages treated with these drugs and infected with S. Typhimurium or Y. pestis led to the identification of potential host-directed mechanisms involved in bacterial clearance. Our observations are striking since we demonstrate that these drugs protect against diverse pathogens via a previously unappreciated mechanism(s) that targets host function and not bacterial growth or virulence determinants. Based on the scientific premise that these drugs modulate central nervous system function to achieve clinical efficacy in patients, we hypothesize that common drug- induced neuroimmune signals may promote a robust host immune response against these pathogens. Indeed we showed that an engineered locked chemokine dimer that effectively recruits neutrophils to the infection site protects mice from such infections. In Aim 1, we will optimize the protective effects of ?lead drugs? and an engineered chemokine singly, in combination, or in conjunction with standard of care antibiotics in animal models of C. difficile and S. Typhimurium infections. In Aim 2, we will evaluate how the ?lead drugs? promote host defenses against bacterial infections and determine how these therapeutics impact neutrophil and macrophage responses from onset to the resolution phase of infection. In Aim 3, we will investigate the relationship between efficacy of the lead drugs and neuroimmune signaling along the microbiota-gut-brain axis, whether these lead candidates alter composition of the microbiota prior to or after C. difficile and S. Typhimurium infections, and characterize the effect of the microbiota composition on the pharmacokinetics and pharmacodynamics of our lead drugs. These studies could lead to broadly active, novel therapeutics against MDR pathogens of immediate concern allowing rapidly moving drug candidates into preclinical and clinical studies for eventual licensure.